Biosensors

11 pages, 7658 KiB

Open AccessArticle

Colorimetric Detection of microRNA-378 Based on Y-Shaped Structure Formed by Gold Nanoparticles and Catalytic Hairpin Self-Assembly

by Yahui Gao, Jinru Pan, Bingyuan Fan, Shan Wang, Qian Wang, Wanru Liu, Fang Hu and Wei Meng

Biosensors 2025, 15(5), 319; https://doi.org/10.3390/bios15050319 - 15 May 2025

Abstract

The timely and accurate detection of cancer is crucial for preventing disease progression and for the early treatment of confirmed cases. MiRNAs are cancer markers. In this study, a simple miRNA detection method is proposed. Three hairpins were designed based on gold nanoparticles [...] Read more.

The timely and accurate detection of cancer is crucial for preventing disease progression and for the early treatment of confirmed cases. MiRNAs are cancer markers. In this study, a simple miRNA detection method is proposed. Three hairpins were designed based on gold nanoparticles combined with catalytic hairpin assembly nucleic acid amplification technology. The low-pH method was used for rapid coupling, and hairpin H1 was opened by miR-378, triggering the cycle reaction and signal amplification and finally forming a Y-shaped structure, thereby narrowing the distance between gold nanoparticles and achieving colorimetric detection. The absorbance change (A₆₂₀/A₅₂₀) was proportional to the concentration of miR-378 (0.05–5 nM), with a detection limit of 0.05 nM. This method also has an evident detection effect on real samples. HeLa and L-02 cell extracts were analyzed using this method. The former showed no obvious color change, whereas the maximum absorption peak of the latter showed a red shift, and the color changed from red to purple. The minimum number of cells that could be detected using HeLa cells was 500 cells/mL. Full article

(This article belongs to the Special Issue Nano Biosensors and Their Applications for In Vivo/Vitro Diagnosis—2nd Edition)

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12 pages, 2162 KiB

Open AccessArticle

Development of Immunoassays for Foodborne Pathogenic Bacteria Detection Using PolyHRP for Signal Enhancement

by Yijia Zhang, Junkang Pan, Qiyi He, Zhihao Xu, Bruce D. Hammock and Dongyang Li

Biosensors 2025, 15(5), 318; https://doi.org/10.3390/bios15050318 - 15 May 2025

Abstract

The rapid and accurate detection of foodborne pathogens is essential for ensuring food safety. Escherichia coli O157:H7 (E. coli O157:H7) and Salmonella Typhimurium (S. Typhimurium) are major foodborne pathogenic bacteria that pose significant public health risks, highlighting the need for [...] Read more.

The rapid and accurate detection of foodborne pathogens is essential for ensuring food safety. Escherichia coli O157:H7 (E. coli O157:H7) and Salmonella Typhimurium (S. Typhimurium) are major foodborne pathogenic bacteria that pose significant public health risks, highlighting the need for effective detection methods. In this study, highly sensitive double-antibody sandwich-based enzyme-linked immunosorbent assays (ELISAs) were developed for the rapid detection of E. coli O157:H7 and S. Typhimurium, utilizing a streptavidin-polymerized horseradish peroxidase (SA-PolyHRP)-based signal enhancement system. Systematic optimization was performed on key parameters, including the capture antibody concentration, detection antibody, and blocking agent. Compared to the method using SA-HRP, substitution with SA-PolyHRP significantly improved detection sensitivity, achieving limits of detection (LODs) of 1.4 × 10⁴ CFU/mL for E. coli O157:H7 and 6.0 × 10³ CFU/mL for S. Typhimurium, with sensitivity enhancements of 7.86-fold and 1.83-fold, respectively. Specificity tests confirmed no cross-reactivity with non-target or closely related pathogenic strains. The matrix effect was effectively mitigated through 10-fold and 100-fold dilutions for E. coli O157:H7 and S. Typhimurium, respectively. Both pathogens were successfully detected in beef samples spiked with 5 CFU after 5 h of incubation. This study demonstrates the effectiveness of PolyHRP-based signal enhancement for the highly sensitive and specific detection of foodborne pathogens, offering a promising approach for rapid food safety monitoring and public health protection. Full article

(This article belongs to the Special Issue Biosensors for Environmental Monitoring and Food Safety)

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11 pages, 2145 KiB

Open AccessArticle

One-Pot Detection of miRNA by Dual Rolling Circle Amplification at Ambient Temperature with High Specificity and Sensitivity

by Wenhua Sun, Kunling Hu, Ziting Song, Ran An and Xingguo Liang

Biosensors 2025, 15(5), 317; https://doi.org/10.3390/bios15050317 - 15 May 2025

Abstract

Rolling circle amplification (RCA) at ambient temperature is prone to false positive signals during nucleic acid detection, which makes it challenging to establish an efficient RCA detection method. The false positive signals are primarily caused by binding of non-target nucleic acids to the [...] Read more.

Rolling circle amplification (RCA) at ambient temperature is prone to false positive signals during nucleic acid detection, which makes it challenging to establish an efficient RCA detection method. The false positive signals are primarily caused by binding of non-target nucleic acids to the circular single-stranded template, leading to non-specific amplification. Here, we present an RCA method for miRNA detection at 37 °C using two circular ssDNAs, each of which is formed by ligating the intramolecularly formed nick (without any splint) in a secondary structure. The specific target recognition is realized by utilizing low concentrations (0.1 nM) of circular ssDNA1 (C1). A phosphorothioate modification is present at G*AATTC on C1 to generate a nick for primer extension during the primer self-generated rolling circle amplification (PG-RCA). The fragmented amplification products are used as primers for the following RCA that serves as signal amplification using circular ssDNA2 (C2). Notably, the absence of splints and the low concentration of C1 significantly inhibits non-target binding, thus minimizing false positive signals. A high concentration (10 nM) of C2 is used to carry out linear rolling circle amplification (LRCA), which is highly specific. This strategy demonstrates a good linear response to 0.01–100 pM of miRNA with a detection limit of 7.76 fM (miR-155). Moreover, it can distinguish single-nucleotide mismatch in the target miRNA, enabling the rapid one-pot detection of miRNA at 37 °C. Accordingly, this method performs with high specificity and sensitivity. This approach is suitable for clinical serum sample analysis and offers a strategy for developing specific biosensors and diagnostic tools. Full article

(This article belongs to the Section Biosensors and Healthcare)

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13 pages, 3541 KiB

Open AccessArticle

Ultrasensitive Bead-Based Immunoassay for Real-Time Continuous Sample Flow Analysis

by Yuri M. Shlyapnikov and Elena A. Shlyapnikova

Biosensors 2025, 15(5), 316; https://doi.org/10.3390/bios15050316 - 15 May 2025

Abstract

The performance of heterophase immunoassays is often limited by the kinetics of analyte binding. This problem is partially solved by bead-based assays, which are characterized by rapid diffusion in the particle suspension. However, at low analyte concentrations, the binding rate is still low. [...] Read more.

The performance of heterophase immunoassays is often limited by the kinetics of analyte binding. This problem is partially solved by bead-based assays, which are characterized by rapid diffusion in the particle suspension. However, at low analyte concentrations, the binding rate is still low. Here, we demonstrate a further improvement of analyte binding kinetics in bead-based immunoassays by simultaneously concentrating both an analyte and magnetic beads in a compact spatial region where binding occurs. The analyte is electrophoretically concentrated in a flow cell where beads are magnetically retained and dragged along the channel by viscous force. The flow cell is integrated with a microarray-based signal detection module, where beads with bound analyte scan the microarray surface and are retained on it by single specific interactions, assuring ultra-high sensitivity of the method. Thus, a continuous flow assay system is formed. Its performance is demonstrated by simultaneous detection of model pathogen biomarkers, cholera toxin (CT) and staphylococcal enterotoxin B (SEB), with a detection limit of 0.1 fM and response time of under 10 min. The assay is capable of real-time online sample monitoring, as shown by a 12 h long continuous flow analysis of tap water for SEB and CT. Full article

(This article belongs to the Special Issue Design, Fabrication, and Applications of Microfluidic Devices for Biosensing)

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15 pages, 6161 KiB

Open AccessArticle

Chiral-Dependent Redox Capacitive Biosensor Using Cu-Cys-GSH Nanoparticles for Ultrasensitive H₂O₂ Detection

by Duygu Yilmaz Aydin, Jie Jayne Wu and Jiangang Chen

Biosensors 2025, 15(5), 315; https://doi.org/10.3390/bios15050315 - 14 May 2025

Abstract

Copper-thiolate nanostructures, formed through the self-assembly of cysteine (Cys) and glutathione (GSH) with copper ions, offer a versatile platform for redox-active applications due to their structural stability and chemical functionality. In this study, Cu-Cys-GSH nanoparticles were synthesized and employed to develop a capacitive [...] Read more.

Copper-thiolate nanostructures, formed through the self-assembly of cysteine (Cys) and glutathione (GSH) with copper ions, offer a versatile platform for redox-active applications due to their structural stability and chemical functionality. In this study, Cu-Cys-GSH nanoparticles were synthesized and employed to develop a capacitive biosensor for the ultralow concentration detection of hydrogen peroxide (H₂O₂). The detection mechanism leverages a Fenton-like reaction, where H₂O₂ interacts with Cu-Cys-GSH nanoparticles to generate hydroxyl radicals (·OH) through redox cycling between Cu²⁺ and Cu⁺ ions. These redox processes induce changes in the sensor’s surface charge and dielectric properties, enabling highly sensitive capacitive sensing at gold interdigitated electrodes (IDEs). The influence of chirality on sensing performance was investigated by synthesizing nanoparticles with both L- and D-cysteine enantiomers. Comparative analysis revealed that the stereochemistry of cysteine impacts the catalytic activity and sensor response, with Cu-L-Cys-GSH nanoparticles exhibiting superior performance. Specifically, the biosensor achieved a linear detection range from 1.0 fM to 1.0 pM and demonstrated an ultra-sensitive detection limit of 21.8 aM, outperforming many existing methods for H₂O₂ detection. The sensor’s practical performance was further validated using milk and saliva samples, yielding high recovery rates and confirming its robustness and accuracy for real-world applications. This study offers a disposable, low-cost sensing platform compatible with sustainable healthcare practices and facilitates easy integration into point-of-care diagnostic systems. Full article

(This article belongs to the Special Issue Innovative Biosensing Technologies for Sustainable Healthcare)

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19 pages, 6179 KiB

Open AccessArticle

Automatic Calculation of Average Power in Electroencephalography Signals for Enhanced Detection of Brain Activity and Behavioral Patterns

by Nuphar Avital, Nataniel Shulkin and Dror Malka

Biosensors 2025, 15(5), 314; https://doi.org/10.3390/bios15050314 - 14 May 2025

Abstract

Precise analysis of electroencephalogram (EEG) signals is critical for advancing the understanding of neurological conditions and mapping brain activity. However, accurately visualizing brain regions and behavioral patterns from neural signals remains a significant challenge. The present study proposes a novel methodology for the [...] Read more.

Precise analysis of electroencephalogram (EEG) signals is critical for advancing the understanding of neurological conditions and mapping brain activity. However, accurately visualizing brain regions and behavioral patterns from neural signals remains a significant challenge. The present study proposes a novel methodology for the automated calculation of the average power of EEG signals, with a particular focus on the beta frequency band which is known for its pronounced activity during cognitive tasks such as 2D content engagement. An optimization algorithm is employed to determine the most appropriate digital filter type and order for EEG signal processing, thereby enhancing both signal clarity and interpretability. To validate the proposed methodology, an experiment was conducted with 22 students, during which EEG data were recorded while participants engaged in cognitive tasks. The collected data were processed using MATLAB (version R2023a) and the EEGLAB toolbox (version 2022.1) to evaluate various filters, including finite impulse response (FIR) and infinite impulse response (IIR) Butterworth and IIR Chebyshev filters with a 0.5% passband ripple. Results indicate that the IIR Chebyshev filter, configured with a 0.5% passband ripple and a fourth-order design, outperformed the alternatives by effectively reducing average power while preserving signal fidelity. This optimized filtering approach significantly improves the accuracy of neural signal visualizations, thereby facilitating the creation of detailed brain activity maps. By refining the analysis of EEG signals, the proposed method enhances the detection of specific neural behaviors and deepens the understanding of functional brain regions. Moreover, it bolsters the reliability of real-time brain activity monitoring, potentially advancing neurological diagnostics and insights into cognitive processes. These findings suggest that the technique holds considerable promise for future applications in brain–computer interfaces and advanced neurological assessments, offering a valuable tool for both clinical practice and research exploration. Full article

(This article belongs to the Section Biosensor and Bioelectronic Devices)

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14 pages, 4266 KiB

Open AccessArticle

One-Step Labeling Based on Eu-MOFs to Develop Fluorescence Side-Flow Immunoassay for AFB1 Detection in Corn

by Yinjun Li, Hua Ding, Ziyu Wang, Zewei Luo and Xitian Peng

Biosensors 2025, 15(5), 313; https://doi.org/10.3390/bios15050313 - 14 May 2025

Abstract

Lateral flow immunoassay (LFIA) is a promising tool for rapid detection in the field of agricultural product analysis due to its advantages of cost-effectiveness and operational simplicity. In this work, Eu metal–organic frameworks (MOFs) were introduced to LFIA as a rapid detection method [...] Read more.

Lateral flow immunoassay (LFIA) is a promising tool for rapid detection in the field of agricultural product analysis due to its advantages of cost-effectiveness and operational simplicity. In this work, Eu metal–organic frameworks (MOFs) were introduced to LFIA as a rapid detection method characterized by high stability and low interference. Key research objectives included strong fluorescence, ease of labeling, and the utilization of fluorescent probes. Eu-MOFs were synthesized in one step via the hydrothermal method, exhibiting a fluorescence lifetime of 163 μs and spherical particles with diameters ranging from 250 to 400 nm. These conditions fulfill the characteristics and requirements of LFIA. Eu-MOFs exploit the porous nature of MOFs to mitigate the drawbacks associated with complex crosslinking agents. This enables antibody proteins to be cross-linked merely upon contact, thereby simplifying the detection process. A time-resolved LFIA method was developed utilizing Eu-MOFs for the detection of aflatoxin B1 (AFB1) in corn, achieving a limit of detection (LOD, IC10) of 0.149 ng/mL. The accuracy and reliability of the Eu-MOFs-LFIA method were validated through comparisons with spiked concentrations during spiking and blind sample analyses, with verification conducted using ultra-high-performance liquid chromatography mass spectrometry (UPLC-MS). Furthermore, testing of real samples demonstrated that the Eu-MOFs-LFIA method can effectively facilitate rapid detection of AFB1 in corn. Full article

(This article belongs to the Special Issue Optical Fiber Biochemical and Environmental Sensors)

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22 pages, 3776 KiB

Open AccessReview

Challenges in Adapting Fibre Optic Sensors for Biomedical Applications

by Sahar Karimian, Muhammad Mahmood Ali, Marion McAfee, Waqas Saleem, Dineshbabu Duraibabu, Sanober Farheen Memon and Elfed Lewis

Biosensors 2025, 15(5), 312; https://doi.org/10.3390/bios15050312 - 13 May 2025

Viewed by 221

Abstract

Fibre optic sensors (FOSs) have developed as a transformative technology in healthcare, often offering unparalleled accuracy and sensitivity in monitoring various physiological and biochemical parameters. Their applications range from tracking vital signs to guiding minimally invasive surgeries, enabling advancements in medical diagnostics and [...] Read more.

Fibre optic sensors (FOSs) have developed as a transformative technology in healthcare, often offering unparalleled accuracy and sensitivity in monitoring various physiological and biochemical parameters. Their applications range from tracking vital signs to guiding minimally invasive surgeries, enabling advancements in medical diagnostics and treatment. However, the integration of FOSs into biomedical applications faces numerous challenges. This article describes some challenges for adopting FOSs for biomedical purposes, exploring technical and practical obstacles, and examining innovative solutions. Significant challenges include biocompatibility, miniaturization, addressing signal processing complexities, and meeting regulatory standards. By outlining solutions to the stated challenges, it is intended that this article provides a better understanding of FOS technologies in biomedical settings and their implementation. A broader appreciation of the technology, offered in this article, enhances patient care and improved medical outcomes. Full article

(This article belongs to the Special Issue Feature Papers of Biosensors)

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10 pages, 1735 KiB

Open AccessCommunication

Wearable Humidity Sensor Using Cs₃Cu₂I₅ Metal Halides with Hydroxyl Selective Phase Transition for Breath Monitoring

by Si Hyeok Yang, Lim Kyung Oh, Dong Ho Lee, Donghoon Gwak, Nara Song, Bowon Oh, Na Young Lee, Hongki Kim, Han Seul Kim and Jin Woo Choi

Biosensors 2025, 15(5), 311; https://doi.org/10.3390/bios15050311 - 13 May 2025

Viewed by 141

Abstract

The low-dimensional metal halide Cs₃Cu₂I₅ exhibits unique electrical and chemical properties. Notably, it undergoes a phase transition to CsCu₂I₃ upon exposure to hydroxyl (-OH) gas, resulting in significant changes in its electrical characteristics. In this [...] Read more.

The low-dimensional metal halide Cs₃Cu₂I₅ exhibits unique electrical and chemical properties. Notably, it undergoes a phase transition to CsCu₂I₃ upon exposure to hydroxyl (-OH) gas, resulting in significant changes in its electrical characteristics. In this study, we developed a highly selective semiconductor-based gas sensor utilizing Cs₃Cu₂I₅. The material was synthesized on an Al₂O₃ substrate with carbon electrodes using a solution-based process, enabling gas sensing based on its electrical properties. The sensor was further integrated into an Arduino-based real-time monitoring system for wearable applications. The final system was mounted onto a face mask, enabling the real-time detection of human respiration. This research presents a next-generation sensor platform for real-time respiratory monitoring, demonstrating the potential of Cs₃Cu₂I₅ in advanced wearable bio-gas sensing applications. Full article

(This article belongs to the Special Issue Wearable Biosensors and Health Monitoring)

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13 pages, 2896 KiB

Open AccessArticle

Individually Modified Microneedle Array for Minimal Invasive Multi-Electrolyte Monitoring

by Ketian Yu, Yukun Ma, Yiming Wei, Wanying Chen, Zhen Dai, Yu Cai, Xuesong Ye and Bo Liang

Biosensors 2025, 15(5), 310; https://doi.org/10.3390/bios15050310 - 12 May 2025

Viewed by 135

Abstract

Electrolytes play crucial roles in regulating nerve and muscle functions. Currently, microneedle technology enables real-time electrolyte monitoring through minimally invasive methods. However, due to the small size of microneedles, performing multi-layer modifications on individual microneedles and ensuring the integrity of these layers pose [...] Read more.

Electrolytes play crucial roles in regulating nerve and muscle functions. Currently, microneedle technology enables real-time electrolyte monitoring through minimally invasive methods. However, due to the small size of microneedles, performing multi-layer modifications on individual microneedles and ensuring the integrity of these layers pose significant challenges. Additionally, the puncture efficiency of the electrodes will be affected by the structure of microneedle array integration. To address these issues, we primarily focus on developing a multi-parameter ion monitoring system based on microneedle arrays. By optimizing the surface reconstruction of electrode substrates, the adhesion between the electrode surface and the modification layer was improved, enhancing the stability of the electrodes. Potassium, sodium, and calcium ion-selective electrodes based on microneedles were fabricated, demonstrating good sensitivity and linearity. To tackle the puncture efficiency of microneedle arrays, finite element simulation was employed to investigate the mechanical properties of different structural designs of microneedle arrays during skin insertion. Ultimately, an integrated microneedle array was designed and assembled, and a multi-parameter ion monitoring system was developed, validated through in vitro simulations and in vivo animal experiments. This research provides valuable insights into the development and advancement of minimally invasive, multi-parameter dynamic monitoring technologies in clinical settings. Full article

(This article belongs to the Section Nano- and Micro-Technologies in Biosensors)

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45 pages, 15184 KiB

Open AccessReview

Wearable Electrochemical Glucose Sensors for Fluid Monitoring: Advances and Challenges in Non-Invasive and Minimally Invasive Technologies

by Ming Wang, Junjie Zheng, Ge Zhang, Shiyan Lu and Jinli Zhou

Biosensors 2025, 15(5), 309; https://doi.org/10.3390/bios15050309 - 12 May 2025

Viewed by 347

Abstract

This review highlights the latest developments in wearable electrochemical glucose sensors, focusing on their transition from invasive to non-invasive and minimally invasive designs. We discuss the underlying mechanisms, performance metrics, and practical challenges of these technologies, emphasizing their potential to revolutionize diabetes care. [...] Read more.

This review highlights the latest developments in wearable electrochemical glucose sensors, focusing on their transition from invasive to non-invasive and minimally invasive designs. We discuss the underlying mechanisms, performance metrics, and practical challenges of these technologies, emphasizing their potential to revolutionize diabetes care. Additionally, we explore the motivation behind this review: to provide a comprehensive analysis of emerging sensing platforms, assess their clinical applicability, and identify key research gaps that need addressing to achieve reliable, long-term glucose monitoring. By evaluating electrochemical sensors based on tears, saliva, sweat, urine, and interstitial fluid, this work aims to guide future innovations toward more accessible, accurate, and user-friendly solutions for diabetic patients, ultimately improving their quality of life and disease management outcomes. Full article

(This article belongs to the Special Issue Wearable and Implantable Bioelectronics for Advanced Biosensing and Human Health Monitoring)

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14 pages, 6839 KiB

Open AccessArticle

Analysis of Colorectal Cancer Gene Mutations and Application of Long Blocker Displacement Amplification Technology for High-Throughput Mutation Detection

by Ping Lu, Xinglei Su, Sirui Leong, Xuehao Xiu, Ping Song, Junjie Peng and Yunpei Si

Biosensors 2025, 15(5), 308; https://doi.org/10.3390/bios15050308 - 12 May 2025

Viewed by 144

Abstract

Genetic mutation detection for colorectal cancer (CRC) is crucial for precision diagnosis and treatment, yet current methods often suffer from challenges such as low sensitivity, time consumption, and high costs. In our preliminary bioinformatic analysis of 751 CRC cases from The Cancer Genome [...] Read more.

Genetic mutation detection for colorectal cancer (CRC) is crucial for precision diagnosis and treatment, yet current methods often suffer from challenges such as low sensitivity, time consumption, and high costs. In our preliminary bioinformatic analysis of 751 CRC cases from The Cancer Genome Atlas and 131 Chinese patient samples, APC, TP53, and KRAS were identified as the most frequently mutated genes. Among them, KRAS missense mutations emerged as key diagnostic biomarkers. In this study, we applied a fluorescence-based long block displacement amplification (LBDA) sensing method for the rapid, high-throughput, and cost-effective detection of KRAS genetic mutations. In the LBDA system, SYBR Green dye binds to the amplified double-stranded DNA, generating a fluorescence signal that directly reflects the abundance of mutant types (MTs). This real-time signal output enables the enrichment and sensitive detection of MTs, establishing LBDA as an efficient biosensing platform for KRAS genotyping. Using this technique, a detection limit of 0.08% variant allele frequency was achieved with 20 ng of synthetic DNA input. To evaluate clinical performance, the LBDA method was applied to 118 tissue samples from 59 CRC patients, including tumor and matched peritumoral tissues. For 59 CRC tumor samples, LBDA successfully identified KRAS mutations in 37.29% of cases, closely matching results (42.37%) obtained by next-generation sequencing and achieving 88% sensitivity and 100% specificity. In conclusion, this study presents a rapid and cost-effective mutation detection method based on optical biosensing, offering strong potential for advancing personalized CRC diagnosis and treatment. Full article

(This article belongs to the Special Issue DNA Molecular Engineering-Based Biosensors)

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14 pages, 2078 KiB

Open AccessArticle

A Microfluidic Device Integrating a Glucose Sensor and Calibration Function for Cell-Based Assays

by Laner Chen, Kenta Shinha, Hiroko Nakamura, Kikuo Komori and Hiroshi Kimura

Biosensors 2025, 15(5), 307; https://doi.org/10.3390/bios15050307 - 11 May 2025

Viewed by 252

Abstract

Microphysiological systems (MPS) incorporating microfluidic technologies offer improved physiological relevance and real-time analysis for cell-based assays, but often lack non-invasive monitoring capabilities. Addressing this gap, we developed a microfluidic cell-based assay platform integrating an electrochemical biosensor for real-time, non-invasive monitoring of kinetic cell [...] Read more.

Microphysiological systems (MPS) incorporating microfluidic technologies offer improved physiological relevance and real-time analysis for cell-based assays, but often lack non-invasive monitoring capabilities. Addressing this gap, we developed a microfluidic cell-based assay platform integrating an electrochemical biosensor for real-time, non-invasive monitoring of kinetic cell status through glucose consumption. The platform addresses the critical limitations of traditional cell assays, which typically rely on invasive, discontinuous methods. By combining enzyme-modified platinum electrodes within a microfluidic device, our biosensor can quantify dynamic changes in glucose concentration resulting from cellular metabolism. We have integrated a calibration function that corrects sensor drift, ensuring accurate and prolonged short-term measurement stability. In the validation experiments, the system successfully monitored glucose levels continuously for 20 h, demonstrating robust sensor performance and reliable glucose concentration predictions. Furthermore, in the cell toxicity assays using HepG2 cells exposed to varying concentrations of paraquat, the platform detected changes in glucose consumption, effectively quantifying the cellular toxicity responses. This capability highlights the device’s potential for accurately assessing the dynamic physiological conditions of the cells. Overall, our integrated platform significantly enhances cell-based assays by enabling continuous, quantitative, and non-destructive analysis, positioning it as a valuable tool for future drug development and biomedical research. Full article

(This article belongs to the Special Issue Microfluidics for Biomedical Applications (3rd Edition))

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19 pages, 6630 KiB

Open AccessArticle

Improving the Accuracy of a Wearable Uroflowmeter for Incontinence Monitoring Under Dynamic Conditions: Leveraging Machine Learning Methods

by Faezeh Shanehsazzadeh, John O. L. DeLancey and James A. Ashton-Miller

Biosensors 2025, 15(5), 306; https://doi.org/10.3390/bios15050306 - 11 May 2025

Viewed by 229

Abstract

Urinary incontinence affects many women, yet there are no monitoring devices capable of accurately capturing flow dynamics during everyday activities. Building on our initial development of a wearable personal uroflowmeter, this study enhances the device’s performance under realistic, dynamic conditions similar to those [...] Read more.

Urinary incontinence affects many women, yet there are no monitoring devices capable of accurately capturing flow dynamics during everyday activities. Building on our initial development of a wearable personal uroflowmeter, this study enhances the device’s performance under realistic, dynamic conditions similar to those encountered in daily living. We integrated an optimized eight-vane Etoile flow conditioner with a 0.2D opening into the device. Both computational fluid dynamics simulations and experimental tests demonstrated that this flow conditioner significantly reduced turbulence intensity by 82% and stabilized the axial velocity profile by 67%, increasing the R² of flow rate measurements from 0.44 to 0.92. Furthermore, our machine learning framework—utilizing a support vector machine (SVM) and an extreme gradient boosting (XGBoost) model with principal component analysis (PCA)—accurately predicted the true flow rate with high correlations, robust performance, and minimal overfitting. For the test dataset, the SVM achieved a correlation of 0.86, an R² of 0.74, and an MAE of 2.8, whereas the XGBoost-PCA model exhibited slightly stronger performance, with a correlation of 0.88, an R² of 0.76, and an MAE of 2.6. These advances established a solid foundation for developing a reliable, wearable uroflowmeter capable of effectively monitoring urinary incontinence in real-world settings. Full article

(This article belongs to the Special Issue Advances in Flexible and Wearable Biosensors)

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21 pages, 5217 KiB

Open AccessArticle

Gait Phase Recognition in Multi-Task Scenarios Based on sEMG Signals

by Xin Shi, Xiaheng Zhang, Pengjie Qin, Liangwen Huang, Yaqin Zhu and Zixiang Yang

Biosensors 2025, 15(5), 305; https://doi.org/10.3390/bios15050305 - 10 May 2025

Viewed by 127

Abstract

In the human–exoskeleton interaction process, accurately recognizing gait phases is crucial for effectively assessing the assistance provided by the exoskeleton. However, due to the similarity in muscle activation patterns between adjacent gait phases, the recognition accuracy is often low, which can easily lead [...] Read more.

In the human–exoskeleton interaction process, accurately recognizing gait phases is crucial for effectively assessing the assistance provided by the exoskeleton. However, due to the similarity in muscle activation patterns between adjacent gait phases, the recognition accuracy is often low, which can easily lead to confusion in surface electromyography (sEMG) feature extraction. This paper proposes a real-time recognition method based on multi-scale fuzzy approximate root mean entropy (MFAREn) and an Efficient Multi-Scale Attention Convolutional Neural Network (EMACNN), building upon the concept of fuzzy approximate entropy. MFAREn is used to extract the dynamic complexity and energy intensity features of sEMG signals, serving as the input matrix for EMACNN to achieve fast and accurate gait phase recognition. This study collected sEMG signals from 10 subjects performing continuous lower limb gait movements in five common motion scenarios for experimental validation. The results show that the proposed method achieves an average recognition accuracy of 95.72%, outperforming the other comparison methods. The method proposed in this paper is significantly different compared to other methods (p < 0.001). Notably, the recognition accuracy for walking in level walking, stairs ascending, and ramp ascending exceeds 95.5%. This method demonstrates a high recognition accuracy, enabling sEMG-based gait phase recognition and meeting the requirements for effective human–exoskeleton interaction. Full article

(This article belongs to the Section Wearable Biosensors)

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16 pages, 1919 KiB

Open AccessArticle

Multi-Parametric Electrochemical Sensing Platform: Applications in Animal Welfare

by C. Ferreira, E. Lynch, A. O’Herlihy, F. Barry, L. C. Nagle, S. R. Teixeira and P. Galvin

Biosensors 2025, 15(5), 304; https://doi.org/10.3390/bios15050304 - 10 May 2025

Viewed by 186

Abstract

The rapid growth of the dairy sector requires advanced monitoring tools to ensure sustainable practices that benefit the environment, economy, and human health. Current monitoring devices often lack multi-parametric capabilities, limiting their ability to provide comprehensive data on critical chemical and biochemical parameters. [...] Read more.

The rapid growth of the dairy sector requires advanced monitoring tools to ensure sustainable practices that benefit the environment, economy, and human health. Current monitoring devices often lack multi-parametric capabilities, limiting their ability to provide comprehensive data on critical chemical and biochemical parameters. To address this challenge, this work presented the integration of a real-time multi-parametric device with sensors for pH, temperature, nitrate, and nitrite, providing a comprehensive solution to dairy cattle health monitoring. This solution included an electrochemical platform, Portable Unit for Lab-on-Site Electrochemistry (PULSE), and an application for data processing and display. In-house fabricated flexible gold-printed electrodes demonstrated accurate detection of nitrite and nitrate when integrated with the PULSE, achieving sensitivities of 6.32

μ

A/ppm/

{cm}^{2}

in artificial interstitial fluid and 1.92

μ

A/ppm/

{cm}^{2}

in phosphate buffered saline, respectively. The PULSE achieved 65.83% and 58.3% lower limits of detection in phosphate buffered saline than a benchtop potentiostat, for nitrate and nitrite, respectively, along with a 24.5% increase in nitrite sensitivity, enhancing its ability to detect lower analyte concentrations. pH sensing was carried out with a commercial screen-printed electrode coated with a layer of iridium oxide. The pH was tested in ruminal complex fluid, obtaining a pH sensitivity of −59.63 mV/pH and an accuracy of 98.9%. These findings highlighted the potential of this technology as an effective tool for dairy cattle health monitoring and its deployment in real-world scenarios. Full article

(This article belongs to the Special Issue Novel Nanomaterials and Nanotechnology: From Fabrication Methods and Improvement Strategies to Applications in Biosensing and Biomedicine (2nd Edition))

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16 pages, 4093 KiB

Open AccessArticle

AFM-Based Monitoring of Enzymatic Activity of Individual Molecules of Cytochrome CYP102A1

by Yuri D. Ivanov, Natalia S. Bukharina, Ivan D. Shumov, Oleg N. Afonin, Vadim Y. Tatur, Anna V. Grudo and Alexander I. Archakov

Biosensors 2025, 15(5), 303; https://doi.org/10.3390/bios15050303 - 10 May 2025

Viewed by 206

Abstract

Herein, we report the use of a nanotechnology-based approach for the study of enzyme-functionalized mica surfaces. Atomic force microscopy (AFM) has been employed for the determination of the catalytic activity of single molecules of heme-containing cytochrome P450 CYP102A1 (CYP102A1) enzyme, which was immobilized [...] Read more.

Herein, we report the use of a nanotechnology-based approach for the study of enzyme-functionalized mica surfaces. Atomic force microscopy (AFM) has been employed for the determination of the catalytic activity of single molecules of heme-containing cytochrome P450 CYP102A1 (CYP102A1) enzyme, which was immobilized on the surface of a mica chip. Height fluctuations in individual molecules of the enzyme were measured under near-native conditions by AFM measurements in liquid using a cantilever with a 10 to 20 nm tip curvature radius. We have found that in the process of enzymatic catalysis, the mean amplitude of height fluctuations in individual enzyme molecules is 1.4-fold higher than that of enzyme molecules in an inactive state. The temperature dependence of the mean amplitude of height fluctuations in cytochrome CYP102A1 has been revealed, and the maximum of this dependence has been observed at 22 °C. The proposed nanotechnology-based approach can be employed in studies of a wide variety of enzymes, which are important for the development of novel diagnostic tests and systems for pharmaceutical analysis. The approach developed in our work will favor further miniaturization of enzyme-based biosensors and the transition from traditional sensors to nanobiosensors. Full article

(This article belongs to the Special Issue Miniature Sensors Based on Highly Efficient Chemical and Biological Sensing Interfaces)

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11 pages, 2977 KiB

Open AccessArticle

An Electrochemical Aptasensor for Accurate and Sensitive Detection of Exosomes Based on Dual-Probe Recognition and Hybridization Chain Reaction

by Haojie Ma, Jie Li, Mengjia Gao, Yan Dong, Yi Luo and Shao Su

Biosensors 2025, 15(5), 302; https://doi.org/10.3390/bios15050302 - 9 May 2025

Viewed by 187

Abstract

The accurate and sensitive detection of tumor-derived exosomes holds significant promise for the early diagnosis of cancer. In this study, an electrochemical aptasensor was developed for the high-performance detection of exosomes by integrating dual-probe recognition and hybridization chain reaction (HCR). A dual-probe recognition [...] Read more.

The accurate and sensitive detection of tumor-derived exosomes holds significant promise for the early diagnosis of cancer. In this study, an electrochemical aptasensor was developed for the high-performance detection of exosomes by integrating dual-probe recognition and hybridization chain reaction (HCR). A dual-probe recognition unit composed of a MUC1 aptamer (MUC1-Apt) probe and cholesterol probe was designed for capturing target exosomes and reducing the interference from free proteins, significantly improving the accuracy of exosome detection. It should be noted that the dual-probe recognition unit was formed in conjunction with the HCR. Moreover, a large number of biotins were also assembled on the HCR product, which were used to capture avidin–horseradish peroxidase (SA-HRP) for signal amplification. The CD63 aptamer (CD63-Apt) was immobilized on the surface of a gold electrode for specifically capturing exosomes to construct a classical sandwiched structure. The loaded SA-HRP can efficiently catalyze the reaction of 3, 3′, 5, 5′ tetramethylbenzidine (TMB) and hydrogen peroxide (H₂O₂) to generate a large electrochemical signal. According to this phenomenon, a linear relationship of this proposed aptasensor was achieved between the electrochemical response and 1 × 10²–1 × 10⁷ particles/mL exosomes, with a detection limit of 45 particles/mL. Moreover, the aptasensor exhibited accepted stability and potential clinical applicability. All results proved that this aptasensor has a promising application in exosome-based disease diagnostics. Full article

(This article belongs to the Special Issue Electrochemical Biosensing Platforms for Food, Drug and Health Safety)

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13 pages, 5541 KiB

Open AccessArticle

Bridging Blood and Skin: Biomarker Profiling in Dermal Interstitial Fluid (dISF) for Minimally Invasive Diagnostics

by Yann Sprunger, Johan Longo, Ali Saeidi and Adrian M. Ionescu

Biosensors 2025, 15(5), 301; https://doi.org/10.3390/bios15050301 - 9 May 2025

Viewed by 248

Abstract

Understanding the biochemical relationship between serum and dermal interstitial fluid (dISF) is critical for advancing minimally invasive diagnostics with wearables and point of care devices focusing on most relevant biomarkers accessible in the ISF. This work compares the composition of dISF and serum [...] Read more.

Understanding the biochemical relationship between serum and dermal interstitial fluid (dISF) is critical for advancing minimally invasive diagnostics with wearables and point of care devices focusing on most relevant biomarkers accessible in the ISF. This work compares the composition of dISF and serum using Xsensio’s microneedle-based collector, which yields an average of 3.4

μ

L/h. In the first study, total protein content, human serum albumin (HSA), and immunoglobulin G (IgG) are quantified in twelve volunteers. A second study is dedicated to screening 50 inflammation-related protein biomarkers across twenty volunteers. The results demonstrate that dISF closely resembles serum in its major protein constituents but at reduced concentrations (e.g., 57% for total protein). Strong correlations are observed between dISF and serum for CRP and SAA (

R^{2} > 0.87

), primarily driven by a subject with pathological levels, demonstrating the ability of dISF to reflect systemic inflammation. This study originally reports NT-proBNP detection at comparable levels in both fluids, suggesting that dISF could serve as a reliable proxy for blood NT-proBNP in the non-invasive diagnosis of cardiac failure. Cytokine profiling reveals 36 detectable cytokines, including several unique to dISF. Notably, interleukin concentrations are found to be highly similar between the two fluids. These experimental findings support dISF as a promising diagnostic medium for monitoring both localized and systemic biomarkers in clinical applications. Full article

(This article belongs to the Special Issue Nano Biosensors and Their Applications for In Vivo/Vitro Diagnosis—2nd Edition)

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17 pages, 2642 KiB

Open AccessArticle

Optimizing Whole-Cell Biosensors for the Early Detection of Crop Infections: A Proof-of-Concept Study

by Nadav Zanger and Evgeni Eltzov

Biosensors 2025, 15(5), 300; https://doi.org/10.3390/bios15050300 - 8 May 2025

Viewed by 175

Abstract

This study presents a proof-of-concept evaluation of optimized whole-cell biosensors designed for the real-time detection of crop infections. Genetically engineered luminescent bacterial strains were used to detect volatile organic compounds (VOCs) emitted by crops during spoilage. Key factors investigated include bacterial uniformity, nutrient [...] Read more.

This study presents a proof-of-concept evaluation of optimized whole-cell biosensors designed for the real-time detection of crop infections. Genetically engineered luminescent bacterial strains were used to detect volatile organic compounds (VOCs) emitted by crops during spoilage. Key factors investigated include bacterial uniformity, nutrient supply, and temperature effects. The results demonstrated that lower temperatures (+4 °C) yielded higher sensor sensitivity and prolonged bacterial viability. A proof-of-concept evaluation was conducted in storage-like conditions, showing effective infection detection in potatoes. These findings underscore the potential of whole-cell-based biosensors for monitoring postharvest production in cold storage environments. Full article

(This article belongs to the Section Biosensor and Bioelectronic Devices)

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17 pages, 13507 KiB

Open AccessArticle

Molecular Association Assay Systems for Imaging Protein–Protein Interactions in Mammalian Cells

by Sung-Bae Kim, Tadaomi Furuta, Suresh Thangudu, Arutselvan Natarajan and Ramasamy Paulmurugan

Biosensors 2025, 15(5), 299; https://doi.org/10.3390/bios15050299 - 8 May 2025

Viewed by 217

Abstract

Molecular imaging probes play a pivotal role in assaying molecular events in various physiological systems. In this study, we demonstrate a new genre of bioluminescent probes for imaging protein–protein interactions (PPIs) in mammalian cells, named the molecular association assay (MAA) probe. The MAA [...] Read more.

Molecular imaging probes play a pivotal role in assaying molecular events in various physiological systems. In this study, we demonstrate a new genre of bioluminescent probes for imaging protein–protein interactions (PPIs) in mammalian cells, named the molecular association assay (MAA) probe. The MAA probe is designed to be as simple as a full-length marine luciferase fused to a protein of interest with a flexible linker. This simple fusion protein alone surprisingly works by recognizing a specific ligand, interacting with a counterpart protein of the PPI, and developing bioluminescence (BL) in mammalian cells. We made use of an artificial intelligence (AI) tool to simulate the binding modes and working mechanisms. Our AlphaFold-based analysis on the binding mode suggests that the hinge region of the MAA probe is flexible before ligand binding but becomes stiff after ligand binding and protein association. The sensorial properties of representative MAA probes, FRB-ALuc23 and FRB-R86SG, are characterized with respect to the quantitative feature, BL spectrum, and in vivo tumor imaging using xenografted mice. Our AI-based simulation of the working mechanisms reveals that the association of MAA probes with the other proteins works in a way to facilitate the substrate’s access to the active sites of the luciferase (ALuc23 or R86SG). We prove that the concept of MAA is generally applicable to other examples, such as the ALuc16- or R86SG-fused estrogen receptor ligand-binding domain (ER LBD). Considering the versatility of this conceptionally unique and distinctive molecular imaging probe compared to conventional ones, we are expecting the widespread application of these probes as a new imaging repertoire to determine PPIs in living organisms. Full article

(This article belongs to the Special Issue AI-Enabled Biosensor Technologies for Boosting Medical Applications)

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16 pages, 4866 KiB

Open AccessArticle

Centrifugation-Induced Stable Colloidal Silver Nanoparticle Aggregates for Reproducible Surface-Enhanced Raman Scattering Detection

by Tianyu Zhou and Zhiyang Zhang

Biosensors 2025, 15(5), 298; https://doi.org/10.3390/bios15050298 - 8 May 2025

Viewed by 241

Abstract

Colloidal noble metal nanoparticle aggregates have demonstrated significant advantages in surface-enhanced Raman scattering (SERS) analysis, particularly for online detection, due to their excellent optical properties, spatial homogeneity, and fluidic compatibility. However, conventional chemically induced aggregation methods (such as salt-induced nanoparticle aggregation) suffer from [...] Read more.

Colloidal noble metal nanoparticle aggregates have demonstrated significant advantages in surface-enhanced Raman scattering (SERS) analysis, particularly for online detection, due to their excellent optical properties, spatial homogeneity, and fluidic compatibility. However, conventional chemically induced aggregation methods (such as salt-induced nanoparticle aggregation) suffer from uncontrolled aggregation, limited stability, and narrow detection windows, which restrict their quantitative and long-term applications. In this study, we developed a non-chemical method for fabricating stable colloidal aggregates from uniform β-cyclodextrin-stabilized silver nanoparticles (β-CD@AgNPs) via centrifugation. By precisely controlling the addition rate of silver nitrate, we synthesized β-cyclodextrin-stabilized silver nanoparticles with a uniform size. Surprisingly, these nanoparticles can form highly dispersed and homogeneous colloidal aggregates simply via centrifugation, which is completely different from the behavior of traditional ligand-modified nanoparticles. Notably, the resulting aggregates exhibit excellent SERS enhancement, enabling the sensitive detection of various dyes at nanomolar levels. Furthermore, they maintain a stable SERS signal (RSD = 6.99%) over a detection window exceeding 1 h, markedly improving signal stability and reproducibility compared with salt-induced aggregates. Additionally, using pyocyanin as a model analyte, we evaluated the quantitative performance of these aggregates (LOD = 0.2 nM), achieving satisfactory recovery (82–117%) in spiked samples of drinking water, lake water, and tap water. This study provides a facile strategy for fabricating stable colloidal SERS substrates and paves the way for the advancement of SERS applications in analytical sciences. Full article

(This article belongs to the Special Issue Miniature Sensors Based on Highly Efficient Chemical and Biological Sensing Interfaces)

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45 pages, 8234 KiB

Open AccessReview

Review of Non-Invasive Imaging Technologies for Cutaneous Melanoma

by Luke Horton, Joseph W. Fakhoury, Rayyan Manwar, Ali Rajabi-Estarabadi, Dilara Turk, Sean O’Leary, Audrey Fotouhi, Steven Daveluy, Manu Jain, Keyvan Nouri, Darius Mehregan and Kamran Avanaki

Biosensors 2025, 15(5), 297; https://doi.org/10.3390/bios15050297 - 7 May 2025

Viewed by 406

Abstract

Imaging technologies are constantly being developed to improve not only melanoma diagnosis, but also staging, treatment planning, and disease progression. We start with a description of how melanoma is characterized using histology, and then continue by discussing nearly two dozen different technologies, including [...] Read more.

Imaging technologies are constantly being developed to improve not only melanoma diagnosis, but also staging, treatment planning, and disease progression. We start with a description of how melanoma is characterized using histology, and then continue by discussing nearly two dozen different technologies, including systems currently used in medical practice and those in development. For each technology, we describe its method of operation, how it is or would be projected to be most commonly used in diagnosing and managing melanoma, and for systems in current use, we identify at least one current manufacturer. We also provide a table including the biomarkers identified by and main limitations associated with each technology and conclude by offering suggestions on specific characteristics that might best enhance a technology’s potential for widespread clinical adoption. Full article

(This article belongs to the Special Issue Advanced Optical Methods for Biosensing)

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23 pages, 5249 KiB

Open AccessReview

Carbon Nanotube-Based Field-Effect Transistor Biosensors for Biomedical Applications: Decadal Developments and Advancements (2016–2025)

by Joydip Sengupta and Chaudhery Mustansar Hussain

Biosensors 2025, 15(5), 296; https://doi.org/10.3390/bios15050296 - 7 May 2025

Viewed by 139

Abstract

Advancements in carbon nanotube-based FET (CNT-FET) biosensors from 2016 to 2025 have boosted their sensitivity, specificity, and rapid detection performance for biomedical purposes. This review highlights key innovations in transducer materials, functionalization strategies, and device architectures, including floating-gate CNT-FETs for detecting cancer biomarkers, [...] Read more.

Advancements in carbon nanotube-based FET (CNT-FET) biosensors from 2016 to 2025 have boosted their sensitivity, specificity, and rapid detection performance for biomedical purposes. This review highlights key innovations in transducer materials, functionalization strategies, and device architectures, including floating-gate CNT-FETs for detecting cancer biomarkers, infectious disease antigens, and neurodegenerative disease markers. Novel approaches, such as dual-microfluidic field-effect biosensor (dual-MFB) structures and carboxylated graphene quantum dot (cGQD) coupling, have further expanded their diagnostic potential. Despite significant progress, challenges in scalability, reproducibility, and long-term stability remain. Overall, this work highlights the transformative potential of CNT-FET biosensors while outlining a roadmap for translating laboratory innovations into practical, high-impact biomedical applications. Full article

(This article belongs to the Special Issue Biosensors Based on Transistors)

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13 pages, 1045 KiB

Open AccessArticle

Rapid and Highly Sensitive Detection of Ricin in Biological Fluids Using Optical Modulation Biosensing

by Eliana Levy, Linoy Golani-Zaidie, Shmuel Burg, Efi Makdasi, Ron Alcalay, Reut Falach, Ofir Schuster and Amos Danielli

Biosensors 2025, 15(5), 295; https://doi.org/10.3390/bios15050295 - 6 May 2025

Viewed by 255

Abstract

Ricin, a highly toxic glycoprotein derived from the seeds of Ricinus communis, poses significant risks in bioterrorism and toxicology due to its rapid absorption and ease of dissemination. Rapid, ultra-sensitive detection is crucial for timely medical intervention and implementing security measures. However, existing [...] Read more.

Ricin, a highly toxic glycoprotein derived from the seeds of Ricinus communis, poses significant risks in bioterrorism and toxicology due to its rapid absorption and ease of dissemination. Rapid, ultra-sensitive detection is crucial for timely medical intervention and implementing security measures. However, existing methods often lack sufficient sensitivity or require lengthy processing, limiting their utility for trigger-to-treat scenarios. Here, we present an optical modulation biosensing (OMB)-based ricin assay capable of detecting low concentrations of ricin in buffer, plasma, and biological samples. The assay combines magnetic-bead-based target capture with fluorescent signal enhancement, achieving a limit of detection (LoD) of 15 pg/mL in buffer and 62 pg/mL in plasma, with a 4-log dynamic range. Optimized protocols reduced the assay time to 60 min, maintaining an LoD of 114 pg/mL in plasma while preserving accuracy and reproducibility. The assay successfully detected ricin in bronchoalveolar lavage fluid and serum from mice that were intranasally exposed to ricin, with signals persisting up to 48 h post exposure. Its rapid, high-throughput capabilities and simplified workflow make the OMB-based assay a powerful tool for toxicology, forensic analysis, and counter-bioterrorism. This study highlights the OMB platform’s potential as a sensitive and robust diagnostic tool for detecting hazardous biological agents. Full article

(This article belongs to the Special Issue Optical Bioimaging and Biosensing)

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18 pages, 1686 KiB

Open AccessArticle

An Extracellular Vesicle (EV) Paper Strip for Rapid and Convenient Estimation of EV Concentration

by Gisela Ströhle, Rebecca Goodrum and Huiyan Li

Biosensors 2025, 15(5), 294; https://doi.org/10.3390/bios15050294 - 6 May 2025

Viewed by 136

Abstract

Extracellular vesicles (EVs) have emerged as promising biomarkers and therapeutic agents, yet their quantification remains technically challenging due to the limitations of conventional methods. Here, a low-cost, fluorescence-based, paper-strip immunoassay is presented for rapid and semi-quantitative estimation of EV concentration, inspired by pH [...] Read more.

Extracellular vesicles (EVs) have emerged as promising biomarkers and therapeutic agents, yet their quantification remains technically challenging due to the limitations of conventional methods. Here, a low-cost, fluorescence-based, paper-strip immunoassay is presented for rapid and semi-quantitative estimation of EV concentration, inspired by pH strips. The assay utilizes nitrocellulose membranes functionalized with capture antibodies (anti-CD63, CD9, CD81) and fluorescent dye (ExoBrite™) for EV detection. Systematic optimization of assay parameters—including dye application sequence, incubation time, antibody configuration, and dye concentration—revealed that labeling EVs with dye and incubating on the nitrocellulose paper strips for 20 min yielded the strongest and most reproducible signal. A 200× dilution of ExoBrite™ dye was determined to provide the best balance between sensitivity and specificity. A standard curve generated through twofold serial dilution of EVs from ovarian cancer cell culture medium confirmed a positive, concentration-dependent fluorescence response, establishing a usable dynamic range. Compared to existing technologies, this platform enables fast, simple-to-implement EV quantification using minimal sample volume and equipment. The simplicity and scalability of the method offer strong potential for use in clinical diagnostics and EV research applications. Full article

(This article belongs to the Special Issue Advanced Biosensors for Disease Screening, Monitoring, Diagnosis and Treatment)

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13 pages, 1455 KiB

Open AccessReview

Quantification of Cisplatin Encapsulated in Nanomedicine: An Overview

by Ziwen Zhang, Jiayu Chen, Tao Wen, Hong Deng, Yiyi Zhang, Hua Guo, Hui Chang, Haiyan Xu and Weiqi Zhang

Biosensors 2025, 15(5), 293; https://doi.org/10.3390/bios15050293 - 6 May 2025

Viewed by 289

Abstract

Cisplatin, which kills cancer cells mainly through DNA crosslinking, has been widely used as a first-line chemotherapeutic agent although it also causes severe side effects. To improve anticancer outcomes, various types of cisplatin-based nanomedicines have been developed, either through direct incorporation or coordination [...] Read more.

Cisplatin, which kills cancer cells mainly through DNA crosslinking, has been widely used as a first-line chemotherapeutic agent although it also causes severe side effects. To improve anticancer outcomes, various types of cisplatin-based nanomedicines have been developed, either through direct incorporation or coordination of cisplatin within nanoparticles (NPs). During the formulation and characterization of cisplatin-loaded NPs, quantitative determination of cisplatin is crucial for both clinically used and newly developed NPs. While NPs facilitate cisplatin delivery, the use of different nanomaterials inevitably complicates its determination and increases the cost of quantification. Currently, there is still a significant demand for an accurate, simple, and cost-effective method to determine cisplatin in NPs, which would facilitate the screening and quality control of cisplatin-based nanomedicines. This review aims to discuss the main strategies for quantifying cisplatin, following a summary of the main types of cisplatin-loaded NPs. Application examples of cisplatin determination in NPs are provided, and the key features of each quantification strategy are compared. In addition, NP-based electrochemical sensors are included as an emerging approach for characterizing cisplatin loaded in NPs. Rational selection of an appropriate cisplatin determination method for NPs according to the quantification principle and specific drug-delivery settings is highly recommended. Full article

(This article belongs to the Special Issue Recent Developments in Nanomaterial-Based Electrochemical Biosensors)

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45 pages, 15218 KiB

Open AccessReview

Comprehensive Analysis of Advancement in Optical Biosensing Techniques for Early Detection of Cancerous Cells

by Ayushman Ramola, Amit Kumar Shakya and Arik Bergman

Biosensors 2025, 15(5), 292; https://doi.org/10.3390/bios15050292 - 5 May 2025

Viewed by 335

Abstract

This investigation presents an overview of various optical biosensors utilized for the detection of cancer cells. It covers a comprehensive range of technologies, including surface plasmon resonance

(S P R)

sensors, which exploit changes in refractive index

(R I)

[...] Read more.

This investigation presents an overview of various optical biosensors utilized for the detection of cancer cells. It covers a comprehensive range of technologies, including surface plasmon resonance

(S P R)

sensors, which exploit changes in refractive index

(R I)

at the sensor surface to detect biomolecular interactions. Localized surface plasmon resonance

(L S P R)

sensors offer high sensitivity and versatility in detecting cancer biomarkers. Colorimetric sensors, based on color changes induced via specific biochemical reactions, provide a cost-effective and simple approach to cancer detection. Sensors based on fluorescence work using the light emitted from fluorescent molecules detect cancer-specific targets with specificity and high sensitivity. Photonics and waveguide sensors utilize optical waveguides to detect changes in light propagation, offering real-time and label-free detection of cancer biomarkers. Raman spectroscopy-based sensors utilize surface-enhanced Raman scattering

(S E R S)

to provide molecular fingerprint information for cancer diagnosis. Lastly, fiber optic sensors offer flexibility and miniaturization, making them suitable for in vivo and point-of-care applications in cancer detection. This study provides insights into the principles, applications, and advancements of these optical biosensors in cancer diagnostics, highlighting their potential in improving early detection and patient outcomes. Full article

(This article belongs to the Special Issue Fiber Optic Biosensors: Advancements and Applications)

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12 pages, 3280 KiB

Open AccessArticle

A Dual-Signal Electrochemiluminescence Sensor for Kanamycin Detection Based on a Self-Enhanced Zr MOF and Single Co-Reactant Competition Mechanism

by Yawen Zhu, Xuemei Wang, Zhiyong Yan, Feifei Zhang, Jianfei Xia, Lili Lv and Zonghua Wang

Biosensors 2025, 15(5), 291; https://doi.org/10.3390/bios15050291 - 5 May 2025

Viewed by 186

Abstract

The dual-signal output self-calibration mode reduces the false positive and negative signals of electrochemiluminescence (ECL) aptamer sensors. A competitive dual-signal ECL platform was designed for the ultrasensitive detection of kanamycin (KAN) using a zirconium metal–organic framework (Zr MOF) and Luminol as ECL emitters. [...] Read more.

The dual-signal output self-calibration mode reduces the false positive and negative signals of electrochemiluminescence (ECL) aptamer sensors. A competitive dual-signal ECL platform was designed for the ultrasensitive detection of kanamycin (KAN) using a zirconium metal–organic framework (Zr MOF) and Luminol as ECL emitters. To enhance the ECL efficiency, a co-reactant (polyethyleneimine, PEI) was covalently bound to the Zr MOF to achieve self-enhanced ECL. Based on the selective interaction between KAN and its aptamer, the Luminol/KAN/Zr MOF-PEI “sandwich” structure was immobilized on the electrode surface. The competition for PEI between emitters increased the Luminol ECL signal and decreased the Zr MOF’s ECL signal. The ratio in ECL signals between the two competitive emitters enabled the quantitative analysis of KAN, achieving a detection limit as low as 7.86 × 10⁻⁴ ng/mL. This study elucidated the synergistic mechanism between self-enhanced ECL and ECL competition, offering a novel approach for constructing dual-signal ECL sensors using a single co-reactant. Full article

(This article belongs to the Special Issue Progress in Electrochemiluminescence Biosensors)

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54 pages, 21736 KiB

Open AccessReview

Whole Cells of Microorganisms—A Powerful Bioanalytical Tool for Measuring Integral Parameters of Pollution: A Review

by Maxim Cheliukanov, George Gurkin, Roman Perchikov, Anastasia Medvedeva, Tatyana Lavrova, Tatyana Belousova, Aleksandra Titova, Yulia Plekhanova, Sergei Tarasov, Anna Kharkova, Vyacheslav Arlyapov, Philippe Mandin, Hideaki Nakamura and Anatoly Reshetilov

Biosensors 2025, 15(5), 290; https://doi.org/10.3390/bios15050290 - 4 May 2025

Viewed by 429

Abstract

Microbial biosensors are bioanalytical devices that can measure the toxicity of pollutants or detect specific substances. This is the greatest advantage of microbial biosensors which use whole cells of microorganisms as powerful tools for measuring integral parameters of environmental pollution. This review explores [...] Read more.

Microbial biosensors are bioanalytical devices that can measure the toxicity of pollutants or detect specific substances. This is the greatest advantage of microbial biosensors which use whole cells of microorganisms as powerful tools for measuring integral parameters of environmental pollution. This review explores the core principles of microbial biosensors including biofuel devices, emphasizing their capacity to evaluate biochemical oxygen demand (BOD), toxicity, heavy metals, surfactants, phenols, pesticides, inorganic pollutants, and microbiological contamination. However, practical challenges, such as sensitivity to environmental factors like pH, salinity, and the presence of competing substances, continue to hinder their broader application and long-term stability. The performance of these biosensors is closely tied to both technological advancement and the scientific understanding of biological systems, which influence data interpretation and device optimization. The review further examines cutting-edge developments, including the integration of electroactive biofilms with nanomaterials, molecular biology techniques, and artificial intelligence, all of which significantly enhance biosensor functionality and analytical accuracy. Commercial implementations and improvement strategies are also discussed, providing a comprehensive overview of the state-of-the-art in this field. Overall, this work consolidates recent progress and identifies both the potential and limitations of microbial biosensors, offering valuable insights into their future development for environmental monitoring. Full article

(This article belongs to the Special Issue Microbial Biosensor: From Design to Applications—2nd Edition)

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